RT Journal Article SR Electronic T1 Studies Relating to the Mode of Action of Methotrexate JF Molecular Pharmacology JO Mol Pharmacol FD American Society for Pharmacology and Experimental Therapeutics SP 318 OP 332 VO 5 IS 4 A1 J. BORSA A1 G. F. WHITMORE YR 1969 UL http://molpharm.aspetjournals.org/content/5/4/318.abstract AB The effect of methotrexate on thymidylate synthesis in mammalian cells in vitro has been studied by using the incorporation of deoxyuridine-6-3H into DNA as an assay for thymidylate synthesis. The results are consistent with the interpretation that inside the living cell methotrexate can inhibit thymidylate synthetase directly. In studies on the effects of this drug on thymidylate synthetase activity in cell-free preparations, using both mammalian and bacterial enzyme sources, inhibition of the synthetase was observed. The inhibition yielded Lineweaver-Burk plots that were uncompetitive with respect to deoxyuridylic acid and competitive with respect to tetrahydrofolate. Values of 1.4 (±0.1) x 10-5 M and 4.3 (±1.2) x 10-6 M were measured for Ki,intercept and Ki, slope, respectively. The possible significance of these findings is discussed. ACKNOWLEDGMENTS We thank Drs. W. R. Bruce and J. E. Till for helpful discussions.